Physiological roles of Na+/Ca2+ exchange in Limulus ventral photoreceptors

In previous work we have presented evidence for electrogenic Na+/Ca2+ exchange in Limulus ventral photoreceptors (1989. J. Gen. Physiol. 93:473-492). This article assesses the contributions to photoreceptor physiology from Na+/Ca2+ exchange. Four separate physiological processes were considered: maintenance of resting sensitivity, light-induced excitation, light adaptation, and dark adaptation. (a) Resting sensitivity: reduction of [Na+]o caused a [Ca2+]o-dependent reduction in light sensitivity and a speeding of the time courses of the responses to individual test flashes; this effect was dependent on the final value to which [Na+]o was reduced. The desensitization caused by Na+ reduction was dependent on the initial sensitivity of the photoreceptor; in fully dark-adapted conditions no desensitization was observed; in light-adapted conditions, extensive desensitization was observed. (b) Excitation: Na+ reduction in fully dark-adapted conditions caused a Ca2+o-dependent depolarizing phase in the receptor potential that persisted beyond the stimulus duration and was evoked by a bright adapting flash. (c) Light adaptation: the degree of desensitization induced by a bright adapting flash was Na+o dependent, being larger with lower [Na+]o. Na+ reduction enhanced light adaptation only at intensities brighter than 4 x 10(-6) W/cm2. In addition to being Na+o dependent, light adaptation was Ca2+o dependent, being greater at higher [Ca2+]o. (d) Dark adaptation: the recovery of light sensitivity after adapting illumination was Na+o dependent. Dark adaptation after bright illumination in voltage-clamped and in unclamped conditions was faster in normal-Na+ saline than in reduced Na+ saline. The final sensitivity to which photoreceptors recovered was lower in reduced-Na+ saline when bright adapting illumination was used. The results suggest the involvement of Na+/Ca2+ exchange in each of these physiological processes. Na+/Ca2+ exchange may contribute to these processes by counteracting normal elevations in [Ca2+]i.     

Measurement of cytosolic Ca2+ concentration in Limulus ventral photoreceptors using fluorescent dyes

Several Ca-sensitive fluorescent dyes (fura-2, mag-fura-2 and Calcium Green-5N) were used to measure intracellular calcium ion concentration, Cai, accompanying light-induced excitation of Limulus ventral nerve photoreceptors. A ratiometric procedure was developed for quantification of Calcium Green-5N fluorescence. A mixture of Calcium Green-5N and a Ca-insensitive dye, ANTS, was injected in the cell and the fluorescence intensities of both dyes were used to calculate the spatial average of Cai within the light-sensitive R lobe of the photoreceptor. In dark-adapted photoreceptors, the initial Cai was 0.40 +/- 0.22 microM (SD, n = 7) as measured with fura-2. Cai peaked in the light-sensitive R lobe at 700-900 ms after the onset of an intense measuring light step, when the spatial average of Cai within the R lobe reached 68 +/- 14 and 62 +/- 37 microM (SD, n = 5) as measured with mag- fura-2 and Calcium Green-5N, respectively. The rate of Cai rise was calculated to be approximately 350 microM/s under the measuring conditions. The resting level of Mg2+ was estimated to be 1.9 +/- 0.9 mM, calculated from mag-fura-2 measurements. To investigate the effect of adapting light on the initial Cai level in the R lobe, a 1-min step of 420 nm background light was applied before each measurement. The first significant (P < 0.05) change in the initial level of Cai occurred even at the lowest adapting light intensity, which delivered approximately 3 x 10(3) effective photons/s. The relative sensitivity of the light-adapted photoreceptors was linearly related to the relative Cai on a double log plot with slope between -4.3 and -5.3. We were unable to detect a Cai rise preceding the light-activated receptor potential. The Cai rise, measured with Calcium Green-5N, lagged 14 +/- 5 ms (SD, n = 32) behind the onset of the receptor potential at room temperature in normal ASW. In the absence of extracellular Ca2+ and at 10 degrees C, this lag increased to 44 +/- 12 ms (SD, n = 17).     

A quantitative comparison of the effects of intracellular calcium injection and light adaptation on the photoresponse of Limulus ventral photoreceptors

Calcium ions were iontophoretically injected into ventral photoreceptors of Limulus by passing current between two intracellular pipettes. Changes in sensitivity and photoresponse time course were measured for both light adaptation and Ca++ injection. We found for some photoreceptors that there was no significant difference in the photoresponse time course for desensitization produced by light adaptation or by Ca++ injection. In other photoreceptors, the time delay of photoresponse for Ca++ injection was slightly longer than for light adaptation. The variability of threshold response amplitude and time delay decreases when the photoreceptor is desensitized by either light adaptation or Ca++ injection. The peak amplitude versus log stimulus intensity relationships for controls, light adaptation, and Ca++ injection all could be described very closely by a single template curve shifted along the log intensity axis. A 40- to 50-fold change in sensitivity is associated with a 2-fold change in photoresponse time delay for both light adaptation and Ca++ injection.     

Microvillar components of light adaptation in blowflies

The process of light adaptation in blowfly photoreceptors was analyzed using intracellular recording techniques and double and triple flash stimuli. Adapting flashes of increasing intensity caused a progressive reduction in the excitability of the photoreceptors, which became temporarily suppressed when 3 x 10(6) quanta were absorbed by the cell. This suppression was confirmed by subsequently applying an intense test flash that photoactivated a considerable fraction of the 10(8) visual pigment molecules in the cell. The period of temporary desensitization is referred to as the refractory period. The stimulus intensity to render the receptor cell refractory was found to be independent of the extracellular calcium concentration over a range of 10(-4) and 10(-2) M. During the refractory period (30-40 ms after the adapting flash) the cell appears to be "protected" against further light adaptation since light absorption during this period did not affect the recovery of the cell's excitability. Calculations showed that the number of quantum absorptions necessary to induce receptor refractoriness is just sufficient to photoactivate every microvillus of the rhabdomere. This coincidence led to the hypothesis that the refractoriness of the receptor cells is due to the refractoriness of the individual microvilli. The sensitivity of the receptor cells after relatively weak adapting flashes was reduced considerably more than could be accounted for by the microvilli becoming refractory. A quantitative analysis of these results suggests that a photoactivated microvillus induces a local adaptation over a relatively small area of the rhabdomere around it, which includes several tens of microvilli. After light adaptation with an intense flash, photoactivation of every microvillus by the absorption of a few quanta produced only a small receptor response whereas photoactivation of every rhodopsin molecule in every microvillus produced the maximum response. The excitatory efficiency of the microvilli therefore increases with the number of quanta that are absorbed simultaneously.     

The phosphoinositide signaling cascade is involved in photoreception in the leech Hirudo medicinalis.

The effects of BAPTA, heparin, and neomycin on electrical light responses were studied in the photoreceptors of Hirudo medicinalis. Light activation produces a fast increase in intracellular Ca2+ concentration (Cai) as detected with the fluorescent Ca2+ indicator calcium green-5N. Chelating intracellular calcium by injections of 10 mmol(-1) BAPTA suppresses spontaneous quantum bumps, reduces light sensitivity by more than 2 log(10) units, and substantially increases the latent period of light responses. BAPTA strongly inhibits the plateau phase of responses to long steps of light. Injections of 45-100 mg ml(-1) of heparin act in a similar manner to BAPTA, affecting the latency of the light responses even more. De-N-sulfated heparin, an inactive analog, is almost ineffective at the same concentration compared with heparin. Heparin diminishes the light-induced Cai elevation significantly, whereas de-N-sulfated heparin does not. Intracellular injections of 50-100 mmol l(-1) of the aminoglycoside neomycin, which inhibits phospholipase-C-mediated inositol 1,4,5-trisphosphate formation, acts similar to BAPTA and heparin. Pressure injections of the hydrolysis resistant analog of inositol 1,4,5-trisphosphate, inositol 2,4,5-trisphosphate, strongly depolarize leech photoreceptors and mimic an effect of light adaptation. These results suggest a close similarity between phototransduction mechanisms in leech photoreceptors and existing models for visual transduction in other invertebrate microvillar photoreceptors.     

Membrane current responses of skate photoreceptors

Light-evoked membrane currents were recorded with suction electrodes from the outer segments of individual photoreceptors enzymatically dissociated from the skate retina. The intensity-response relation of dark-adapted cells closely followed a Michaelis function for which a half-saturating response was elicited by a flash intensity that produced about 36 photoisomerizations. Dim-light responses, as well as the early rising phase of the responses to a wide range of flash intensities, could be described by a reaction scheme that involved a series of four first-order delay stages. The number of delay stages required to model the rising phase of the photocurrents did not change in light adaptation. However, background illumination that reduced sensitivity by 1.5 log units, or a bleaching exposure that resulted in a nearly equivalent desensitization, shortened significantly the time scale of the responses. In both instances there were two- to threefold increases in the rate constants of the transitional delays, and almost complete suppression of the tail current that characterized the response of the dark-adapted cell. These findings suggest that although light adaptation alters the gain and kinetics of the transduction mechanism, the nature of the intervening processes is the same in dark- and light-adapted photoreceptors. Moreover, the results show clearly that there is no need to postulate the existence of a second class of cone-like rods to account for the remarkable ability of skate photoreceptors to respond to incremental stimuli presented on "saturating" background fields or after exposure to an intense bleaching light.     

Adaptation in the Ventral Eye of Limulus is Functionally Independent of the Photochemical Cycle, Membrane Potential, and Membrane Resistance

The early receptor potential (ERP), membrane potential, membrane resistance, and sensitivity were measured during light and/or dark adaptation in the ventral eye of Limulus. After a bright flash, the ERP amplitude recovered with a time constant of 100 ms, whereas the sensitivity recovered with an initial time constant of 20 s. When a strong adapting light was turned off, the recovery of membrane potential and of membrane resistance had time-courses similar to each other, and both recovered more rapidly than the sensitivity. The receptor depolarization was compared during dark adaptation after strong illumination and during light adaptation with weaker illumination; at equal sensitivities the cell was more depolarized during light adaptation than during dark adaptation. Finally, the waveforms of responses to flashes were compared during dark adaptation after strong illumination and during light adaptation with weaker illumination. At equal sensitivities (equal amplitude responses for identical flashes), the responses during light adaptation had faster time-courses than the responses during dark adaptation. Thus neither the photochemical cycle nor the membrane potential nor the membrane resistance is related to sensitivity changes during dark adaptation in the photoreceptors of the ventral eye. By elimination, these results imply that there are (unknown) intermediate process(es) responsible for adaptation interposed between the photochemical cycle and the electrical properties of the photoreceptor.     

Calcium regulates some, but not all, aspects of light adaptation in rod photoreceptors

The role of calcium as a regulator of light adaptation in rod photoreceptors was examined by manipulation of the intracellular Ca2+ concentration through the use of the calcium ionophore A23187 and external Ca2+ buffers. These studies utilized suspensions of isolated and purified frog rod outer segments that retain their mitochondria-rich inner segments (OS-IS). Three criteria of the dark- and light-adapted flash response were characterized as a function of the Ca2+ concentration: (a) the time to peak, (b) the rate of recovery, and (c) the response amplitude or sensitivity. For all Ca2+ concentrations examined, the time to peak of the flash response was accelerated in the presence of background illumination, suggesting that mechanisms controlling this aspect of adaptation are independent of the Ca2+ concentration. The recovery kinetics of the flash response appeared to depend on the Ca2+ concentration. In 1 mM Ca2+-Ringer's and 300 nM Ca2+-Ringer's + A23187, background illumination enhanced the recovery rate of the response; however, in 10 and 100 nM Ca2+-Ringer's + A23187, the recovery rates were the same for dark- and light-adapted responses. This result implies that a critical level of Ca2+ may be necessary for background illumination to accelerate the recovery of the flash response. The sensitivity of the flash response in darkness (SDF) was dependent on the Ca2+ concentration. In 1 mM Ca2+-Ringer's SDF was 0.481 pA per bleached rhodopsin (Rh*); a background of four Rh*/s decreased SDF by half (Io). At 300 nM Ca2+ + A23187, SDF was reduced to 0.0307 pA/Rh* and Io increased to 60 Rh*/s. At 100 nM Ca2+ + A23187, SDF was reduced further to 0.0025 pA/Rh* and Io increased to 220 Rh*/s. In 10 nM Ca2+ + A23187, SDF was lowered to 0.00045 pA/Rh* and Io raised to 760 RhI/s. Using these values of SDF and Io for each respective Ca2+ concentration, the dependence of the flash sensitivity on background intensity could be described by the Weber-Fechner relation. Under low Ca2+ conditions + A23187, bright background illumination could desensitize the flash response. These results are consistent with the idea that the concentration of Ca2+ may set the absolute magnitude of response sensitivity in darkness, and that there exist mechanisms capable of adapting the photoresponse in the absence of significant changes in cytoplasmic Ca2+ concentration.     

Calcium measurement in the periphery of an axon

Aequorin was microinjected into squid giant axons, the axons were stimulated, and the change in light emission was followed. This response was compared with that found when the axon, in addition to being microinjected with aequorin, is also injected with the dye phenol red. Large concentrations of phenol red injected into axons result in a high probability that photons emitted by aequorin, when it reacts with Ca in the core of the axoplasm, will be absorbed before they escape from the axon; photons produced by the aequorin reaction at the periphery of the axoplasm are much less likely to be absorbed. This technique thus favors observing changes in Cai taking place in the periphery of the axon. Stimulation in 50 mM Ca seawater of an aequorin-phenol red-injected axon at 180 s-1 for 1 min produces a scarcely detectable change in Cai; the addition of 2 mM cyanide (CN) to the seawater produces an easily measureable increase in Cai, suggesting that mitochondrial buffering in the periphery is substantial. Making the pH of the axoplasm of a normal axon alkaline with 30 mM NH4+ -50 mM Ca seawater, reduces the resting glow of the axon but results in an even more rapid increase in Cai with stimulation. In a phenol red-injected axon, this treatment results in a measureable response to stimulation in the absence of CN.     

Lanthanum mimicks the trp photoreceptor mutant of Drosophila in the blowfly Calliphora

The effect of lanthanum on the light response of blowfly (Calliphora erythrocephala) photoreceptors was studied. The electrophysiological behaviour of the photoreceptors in the presence of La can be summarized as follows: 1. Upon long stimulation the photoreceptors responded with a 'transient receptor potential', i.e. the cells depolarized at the onset of the stimulus and then repolarized to (or below) the resting potential. This effect was dependent on stimulus intensity and occurred only at high intensities. During illumination membrane noise was reduced. 2. The light-induced changes in membrane potential were paralleled by changes in membrane resistance. 3. The time course of the receptor response was slowed down. 4. Light adaptation led to an increase in response latency. 5. The recovery of the receptor response after light adaptation was slowed down. 6. The sensitivity of the receptor cells measured by the response to short light stimuli was reduced. In summary, the electrophysiological behaviour of Calliphora photoreceptors in the presence of La was very similar to that of the photoreceptors of the trp (transient receptor potential) mutant of Drosophila melanogaster. This result suggests that La and trp mutation affect the same cellular processes in the photoreceptors.     

Spectral sensitivity of light induced respiratory activity of photoreceptor mitochondria in the intact fly

Summary FlyCalliphora erythrocephala (white eyed) photoreceptors were investigated in intact, living animals by microspectrofluorometry in vivo. The fluorescence of mitochondrial flavoproteins (Tinbergen and Stavenga 1986) was used to monitor transient changes in oxidative metabolism, which were induced by a test light following a stimulus of variable intensity.Two stimulus types were applied, a brief, activating illumination and a prolonged, adapting illumination, respectively. The intensity ranges of activation and adaptation appear to be separated by ca. 3 log units.Action spectra for inducing a criterion activation or adaptation of the light-dependent mitochondrial system are virtually indistinguishable and closely resemble the spectral sensitivity measured electrophysiologically, thus reinforcing the hypothesis (Hamdorf and Langer 1966; Stavenga and Tinbergen 1983) that the light-induced changes in oxidative metabolism in fly photoreceptors are closely linked to the phototransduction process.On the basis of the literature we conclude that a light-induced rise in cytosolic calcium concentration is the likely cause for enhancing mitochondrial activity.     

Intracellular calcium changes in Balanus photoreceptor. A study with calcium ion-selective electrodes and Arsenazo III

Intracellular Ca2+ concentration (Cai) in the dark and during light stimulation, was measured in Balanus photoreceptors with Ca2+ ion-selective electrodes (Ca-ISE) and Arsenazo III absorbance changes (AIII). The average basal Cai of 17 photoreceptors in darkness was 300 +/- 160 nM determined with liquid ion-exchanger (t-HDOPP) Ca-ISE. Ca-ISE measurements indicated that light increased Cai by 700 nM (average), whereas AIII indicated an average change of 450 nM. The time course of AIII absorbance changes matched the time course of changes in the receptor potential more closely than did the Ca-ISE. Changes in Cai were graded with light intensity but the change in Cai was much greater for a decade change in intensity at high light intensity than at low intensity. The peak light induced conductance change of voltage clamped cells had a relationship to light intensity similar to that of the change in Cai. The peak Cai level measured with Ca-ISE was in good agreement with the free Ca2+ concentration of injected buffer solutions. Control Cai levels were usually restored within 5 min following injection of Ca2+ buffers. Injection of Ca2+ buffers with free Ca2+ of 0.6 microM produced a membrane depolarization. Larger increases in Cai (greater than microM) produced by injection of CaCl2 or release of Ca2+ from injected buffers by acidifying the cell, produced a pronounced membrane hyperpolarization. Increasing Cai with all of these techniques reduced the amplitude of the receptor potential. The time course of the receptor potential recovery was usually similar to that of Cai recovery.     

A lingering elevation of Cai accompanies inhibition of inositol 1,4,5 trisphosphate-induced Ca release in Limulus ventral photoreceptors

Injection of inositol 1,4,5 trisphosphate (InsP3) into Limulus ventral photoreceptors causes an elevation of intracellular free Ca concentration (Cai) and depolarizes the photoreceptors. When measured with the photoprotein aequorin, the InsP3-induced Cai increase follows the time course of depolarization and declines within 1-2 s. However, sensitivity to further injections of InsP3 remains suppressed for several tens of seconds. The possibility that the suppression of Ca release (feedback inhibition) is due to a small lingering elevation of Cai, below the existing detection limit of aequorin, was investigated by measuring Cai with Ca-sensitive electrodes. Double-barreled, Ca- selective microelectrodes were used to pressure inject InsP3 and measure Cai at the same point. Light or InsP3 injections into the light- sensitive compartment depolarized the photoreceptors and induced an elevation of Cai that persisted for tens of seconds. Injections of InsP3 during the decay of Cai showed that sensitivity to InsP3 recovered as resting Cai approached the prestimulus level. The relationship between elevated Cai and feedback inhibition was very steep. An elevation of Cai of 1 microM or more was associated with inhibitions of 79 +/- 12.4% (SEM; n = 7) for the InsP3-induced Cai increase and of 76 +/- 8% for depolarizations. With a residual Cai elevation of 0.01 microM or less, the mean inhibition was 10 +/- 7.4% for InsP3-induced Cai increase and 6.6 +/- 4% for InsP3-induced depolarization. Injections of InsP3 into a light-insensitive compartment within the cell induced elevations of Cai with no associated depolarizations or feedback inhibition. To verify that a sustained elevation of Cai is necessary for inhibition of InsP3-induced Cai increase and depolarization, we injected ethyleneglycol-bis-(beta- aminoethylether)-N,N'-tetraacetic acid (EGTA) between two injections of InsP3. Injection of 1 mM EGTA or the related Ca chelator BAPTA, delivered 750 ms after the first injection of InsP3, restored the peak depolarization caused by the second injection of InsP3 to > 80 +/- 3% of control, compared with 13 +/- 8% without an intervening injection of EGTA. Measurement of Cai with aequorin showed that an intervening injection of EGTA partially restored the InsP3-induced Cai increase. The results suggest that feedback inhibition of InsP3-induced Cai increase and depolarization is mediated by a lingering elevation of Cai and not by depletion of intracellular Ca stores.     

Responses of crayfish photoreceptor cells following intense light adaptation

Summary After intense orange adapting exposures that convert 80% of the rhodopsin in the eye to metarhodopsin, rhabdoms become covered with accessory pigment and appear to lose some microvillar order. Only after a delay of hours or even days is the metarhodopsin replaced by rhodopsin (Cronin and Goldsmith 1984). After 24 h of dark adaptation, when there has been little recovery of visual pigment, the photoreceptor cells have normal resting potentials and input resistances, and the reversal potential of the light response is 10–15 mV (inside positive), unchanged from controls. The log V vs log I curve is shifted about 0.6 log units to the right on the energy axis, quantitatively consistent with the decrease in the probability of quantum catch expected from the lowered concentration of rhodopsin in the rhabdoms. Furthermore, at 24 h the photoreceptors exhibit a broader spectral sensitivity than controls, which is also expected from accumulations of metarhodopsin in the rhabdoms. In three other respects, however, the transduction process appears to be light adapted: (i) The voltage responses are more phasic than those of control photoreceptors. (ii) The relatively larger effect (compared to controls) of low extracellular Ca⁺⁺ (1 mmol/1 EGTA) in potentiating the photoresponses suggests that the photoreceptors may have elevated levels of free cytoplasmic Ca⁺⁺. (iii) The saturating depolarization is only about 30% as large as the maximal receptor potentials of contralateral, dark controls, and by that measure the log V-log I curve is shifted downward by 0.54 log units. The gain (change in conductance per absorbed photon) therefore appears to have been diminished.     

Intracellular calcium transients and developed tension in rat heart muscle. A mechanism for the negative interval-strength relationship

The purposes of the present study were to determine (a) whether changes of intracellular [Ca2+] (Cai) can account for the decrease of developed tension observed in rat heart muscle when stimulation rate is increased, and (b) whether the effect of stimulation rate on Cai is altered in conditions in which the rate of repriming of the sarcoplasmic reticulum (SR) is altered, as when perfusate [Ca2+] (Cao) is increased, and in heart muscle from senescent animals. The photoprotein aequorin was used to monitor Cai in rat papillary muscles. In muscles from 6-mo-old rats, increasing the stimulation rate in the range 0.2-0.66 Hz led to parallel decreases of both the aequorin light transient and developed tension when Cao was 2 mM. When Cao was increased to 4 mM, changes in the stimulation rate had less effect on both the light transient and tension. At 8 mM Cao, changing the stimulation rate had no effect on either the light transient or developed tension. Papillary muscles from 24-mo-old rats, in which SR function is likely to be depressed, exhibited a prolonged Ca2+ transient and twitch. At a Cao of 4 or 8 mM, increasing the stimulation rate from 0.33 to 0.66 Hz still led to decreases in the size of the aequorin light transient and developed tension in these muscles. Developed tension and aequorin light responded to increases of Cao in the same way in both groups of muscles. We conclude that under the conditions of our experiments, developed tension is determined by Cai. The negative interval-strength relationship observed when Cao is in the physiological range can be accounted for by a time-dependent recycling of Ca2+ by the SR. The effects of increasing Cao and the age-related differences observed at high Cao can also be accounted for using this model.     

Evidence for electrogenic Na+/Ca2+ exchange in Limulus ventral photoreceptors

The Ca2+ indicator photoprotein, aequorin, was used to estimate and monitor intracellular Ca2+ levels in Limulus ventral photoreceptors during procedures designed to affect Na+/Ca2+ exchange. Dark levels of [Ca2+]i were estimated at 0.66 +/- 0.09 microM. Removal of extracellular Na+ caused [Ca2+]i to rise transiently from an estimated 0.5-0.6 microM in a typical cell to approximately 21 microM; [Ca2+]i approached a plateau level in 0-Na+ saline of approximately 5.5 microM; restoration of normal [Na+]o lowered [Ca2+]i to baseline with a time course of 1 log10 unit per 9 s. The apparent rate of Nao+-dependent [Ca2+]i decline decreased with decreasing [Ca2+]i. Reintroduction of Ca2+ to 0-Na+, 0-Ca2+ saline in a typical cell caused a transient rise in [Ca2+]i from an estimated 0.36 microM (or lower) to approximately 16.5 microM. This was followed by a decline in [Ca2+]i approaching a plateau of approximately 5 microM; subsequent removal of Cao2+ caused [Ca2+]i to decline slowly (1 log unit in approximately 110 s). Intracellular injection of Na+ in the absence of extracellular Na+ caused a transient rise in [Ca2+]i in the presence of normal [Ca2+]o; in 0-Ca2+ saline, however, no such rise in [Ca2+]i was detected. Under constant voltage clamp (-80 mV) inward currents were measured after the addition of Nao+ to 0-Na+ 0-Ca2+ saline and outward currents were measured after the addition of Cao2+ to 0-Na+ 0-Ca2+ saline. The results suggest the presence of an electrogenic Na+/Ca2+ exchange process in the plasma membrane of Limulus ventral photoreceptors that can operate in forward (Nao+-dependent Ca2+ extrusion) or reverse (Nai+-dependent Ca2+ influx) directions.     

Light-induced extracellular changes of calcium and sodium concentrations in the Planarian ocellus

Ion-selective double-barreled microelectrodes inserted into a planarian ocellus were used to monitor the ocellus potential and the changes in extracellular concentrations of Ca2+ (Ca(o)) and Na+ (Na(o)) caused by a 0.5-sec light flash or sustained (120s) illumination. Ca(o) and Na(o) were slightly decreased following a flash. Sustained illumination caused a biphasic change in Ca(o) (a rapid decrease followed by a slow increase) and a tonic decrease in Na(o). When Na+ in the planarian saline was replaced by Li+ or choline+, the increase in Ca(o) was prevented: sustained illumination induced only a decrease in Ca(o). These results suggest that illumination induces influxes of both Ca+ and Na+ into planarian photoreceptors, and that the Ca2+ influx is rapidly followed by a Na-dependent Ca2+ efflux due to Na-Ca exchange.     

Actions of neomycin on electrical light responses, Ca2+ release, and intracellular Ca2+ changes in photoreceptors of the honeybee drone

Neomycin, known to inhibit phospholipase C-mediated IP3 formation, was applied in the bath or injected into cells and its effects on electrical light responses were analyzed. Neomycin effects on inositol 1,4,5-trisphosphate- and Ca2+-induced Ca2+ release from the endoplasmic reticulum and/or the light-induced Ca2+ elevation were also studied. Neomycin (0.5 mmol x l(-1)) blocked inositol 1,4,5-trisphosphate-, caffeine-, and Ca2+-induced Ca2+ release. Bath application of neomycin decreased the sensitivity to 20-ms light flashes by a factor of up to 100 and slowed the kinetics of dim flash responses. Intracellularly injected neomycin desensitized the photoreceptors more than 1 log unit, increased the latency, and slowed the rate of rise of the light response. Neomycin (0.5 mmol x l(-1)) in the bath delayed and reduced the transient component of responses to 1-s steps of light at intermediate intensities. It also decreased and slowed the light-induced, and it blocked the caffeine-induced intracellular Ca2+ elevation. The combined pharmacological effects of neomycin are suggested to decrease the Ca2+-mediated amplification of the phototransduction cascade and the Ca2+-mediated acceleration of processes determining the kinetics of light responses.     

Light-induced changes in GTP and ATP in frog rod photoreceptors. Comparison with recovery of dark current and light sensitivity during dark adaptation

Light decreases GTP and ATP levels in purified suspensions of physiologically active frog rod outer segments still attached to their inner segment ellipsoids (OS-IS). (a) The GTP decrease is slower in OS-IS (t1/2 = 40 s) than in isolated outer segments (t1/2 = 7 s), which suggests there is more effective buffering in OS-IS. (b) The GTP decrease becomes detectable only at intensities greater than those required to saturate the photoresponse. As the intensity of a continuous light is increased over 4 log units, GTP levels decrease linearly with log intensity by as much as 60%. GTP is reduced to steady intermediate levels during extended illumination of intermediate intensity. (c) At levels of illumination bleaching greater than 0.003% of the rhodopsin, a decrease in ATP levels becomes detectable. (d) Following a flash, GTP levels fall and then rise with a recovery time dependent on the intensity of the flash. (e) After both 0.2 and 2% flash bleaches, the recovery of GTP levels parallels the recovery of light sensitivity, which is slower than the recovery of the dark current. This raises the possibility of a link between GTP levels and light sensitivity.     

Role of intracellular calcium and sodium in light adaptation in the retina of the honey bee drone (Apis mellifera, L)

In the honey bee drone, the decrease in sensitivity to light of a retinula cell exposed to background illumination was found to be accurately reflected by the difference in amplitude between the initial transient depolarization and the lowest steady depolarization evoked by the background light. It is shown that both the decrease in sensitivity to light and the accompanying drop in potential from the transient to the plateau can be prevented by injecting EGTA intracellularly. A decrease in duration and amplitude of responses to short test flashes such as observed immediately after illumination was found to occur too when Ca or Na, but not K, Li, or Mg injected into dark-adapted retinula cells. Injection of EGTA into a retinula cell maintained a steady state of light adaptation, was found to cause an increase in amplitude and duration of the response to a short test flash, thus producing the effects of dark adaptation. It is suggested that, in the retina of the honey bee drone, an increase in intracellular calcium concentration plays a central role in light adaptation and that an increase in intracellular sodium concentration, resulting from the influx of sodium ions during the responses to light, could lead to this increase in intracellular free calcium.